The purpose of the application is to continue and extend cellular neurophysiological studies on opioid peptides and opiates in two brain regions, the hippocampus and the nucleus accumbens, that contain proenkephalin- and prodynorphin-derived peptides. These regions were chosen because of their suggested roles in opiate reinforcement, as cellular models for opiate reward or dependence. The overall objective is to determine the physiological role of central opioid peptide-containing neurons in normal (Naive) subjects and those chronically-treated with opiates, using intracellular (current and voltage-clamp) and whole-cell clamp recording in rat brain slices in vitro. Our hippocampal studies during the last funding period have confirmed a dynorphin-induced augmentation of the M-current, a voltage-dependent K conductance, and have found that nociceptin (orphanin FQ), the agonist for orphan' receptor (ORL1), also augments IM via an opiate receptor mechanism and elicits another non-opiate receptor-mediated action likely to involve a K conductance. In addition, our studies in slices of nucleus accumbens core confirmed the opiate reduction of synaptic transmission in the nucleus accumbens and found pronounced interaction of opiates with glutamate receptor agonists, and especially with those acting on non-NMDA and NMDA receptor-mediated neurotransmission, in neurons from rats treated chronically with morphine. Therefore, our specific aims for the proposed funding period are to: 1) determine the second messenger(s) mediating the dynorphin and nociceptin effects on Im in CA1 hippocampal neurons. 2) Examine the effects of opioid on pharmacologically-isolated EPSCs and responses to NMDA, AMPA and kainate in CA1 of hippocampus. 3) Perform a battery of pharmacological tests to determine the site of action of chronic morphine treatment and possible changes in the postsynaptic NMDA receptor subunit composition in rat nucleus accumbens neurons. We also will use immunohistochemical methods, with selective antibodies to the NMAR2 subunits, and single-cell reverse transcriptase PCR, to determine if chronic morphine treatment causes a cellular and subcellular re- distribution of NMDA receptor subunits. These models will allow later test of several hypotheses, including those concerning neuroadaptative mechanisms following chronic opiate treatment. These studies should help more clearly specify the role of opioid and nociceptin peptides and their receptor in normal, opiate- seeking and opiate addictive-tolerant behavior, and help to provide a cellular basis for therapeusis of opiate craving'.